1. Field of the Invention
The present invention relates to cellulase variants, i.e., endo-beta-1,4-glucanase variants, derived from a parental cellulase, i.e., endo-beta-1,4-glucanase, by substitution, insertion and/or deletion, which variant has a catalytic core domain, in which the variant at position 5 holds an alanine residue (A), a serine residue (S), or a threonine residue (T); at position 8 holds a phenylalanine residue (F), or a tyrosine residue (Y), at position 9 holds a phenylalanine residue (F), a tryptophan residue (W), or a tyrosine residue (Y); at position 10 holds an aspartic acid residue (D), and at position 121 holds an aspartic acid residue (D).
2. Description of Related Art
Cellulases or cellulolytic enzymes are enzymes involved in hydrolysis of cellulose. In the hydrolysis of native cellulose, it is known that there are three major types of cellulase enzymes involved, namely cellobiohydrolase (1,4-beta-D-glucan cellobiohydrolase, EC 3.2.1.91), endo-beta-1,4-glucanase (endo-1,4-beta-D-glucan 4-glucanohydrolase, EC 3.2.1.4) and beta-glucosidase (EC 3.2.1.21).
Especially the endo-beta-1,4-glucanases (EC No. 3.2.1.4) constitute an interesting group of hydrolases for the mentioned industrial uses. Endoglucanases catalyses endo hydrolysis of 1,4-beta-D-glycosidic linkages in cellulose, cellulose derivatives (such as carboxy methyl cellulose and hydroxy ethyl cellulose), lichenin, beta-1,4 bonds in mixed beta-1,3 glucans such as cereal beta-D-glucans or xyloglucans and other plant material containing cellulosic parts. The authorized name is endo-1,4-beta-D-glucan 4-glucano hydrolase, but the abbreviated term endoglucanase is used in the present specification. Reference can be made to T.-M. Enveri, “Microbial Cellulases” in W. M. Fogarty, Microbial Enzymes and Biotechnology, Applied Science Publishers, p. 183-224 (1983); Methods in Enzymology, 1988, Vol. 160, pp. 200-391 (edited by Wood, W. A. and Kellogg, S. T.); Béguin, P., “Molecular Biology of Cellulose Degradation”, Annu. Rev. Microbiol., 1990, Vol. 44, pp. 219-248; Béguin, P. and Aubert, J-P., “The biological degradation of cellulose”, FEMS Microbiology Reviews, 1994, Vol. 13, pp. 25-58; Henrissat, B., “Cellulases and their interaction with cellulose”, Cellulose, 1994, Vol. 1, pp. 169-196.
Cellulases are synthesized by a large number of microorganisms which include fungi, actinomycetes, myxobacteria and true bacteria but also by plants. Especially endoglucanases of a wide variety of specificities have been identified.
A very important industrial use of cellulolytic enzymes is the use for treatment of cellulosic textile or fabric, e.g., as ingredients in detergent compositions or fabric softener compositions, for bio-polishing of new fabric (garment finishing), and for obtaining a “stone washed” look of cellulose-containing fabric, especially denim, and several methods for such treatment have been suggested, e.g., in GB-A-1 368 599, EP-A-0 307 564 and EP-A-0 435 876, WO 91/17243, WO 91/10732, WO 91/17244, PCT/DK95/000108 and PCT/DK95/00132. Another important industrial use of cellulolytic enzymes is the use for treatment of paper pulp, e.g., for improving the drainage or for deinking of recycled paper.
It is also known that cellulases may or may not have a cellulose binding domain (a CBD). The CBD enhances the binding of the enzyme to a cellulose-containing fiber and increases the efficacy of the catalytic active part of the enzyme
Fungi and bacteria produces a spectrum of cellulolytic enzymes (cellulases) which, on the basis of sequence similarities (hydrophobic cluster analysis), can be classified into different families of glycosyl hydrolases [Henrissat B & Bairoch A; Biochem. J., 1993, 293; 781-788]. At present are known cellulases belonging to the families 5, 6, 7, 8, 9, 10, 12, 26, 44, 45, 48, 60, and 61 of glycosyl hydrolases.
Industrially well-performing endo-beta-1,4-glucanases are described in, e.g., WO 91/17243, WO 91/17244 and WO 91/10732, and specific cellulase variants are described in WO 94/07998.
It is an object of the present invention to provide novel variants of cellulolytic enzymes, which variants, when compared to the parental enzyme, show improved performance.